Abstract The larynx is a strategically important organ situated at the crossroads of the upper respiratory and digestive tracts, orchestrating swallowing, breathing, coughing and, in humans, voice. The hallmark of many clinically important laryngeal diseases affecting these critical functions is mucosal inflammation. In other parts of the body, the pathogenesis of mucosal inflammatory disease is intimately linked to local immune responses. The long-term goal of our research is to improve prevention and management of vocal fold disease by identifying and manipulating the manner in which epithelial cell-fibroblast interactions modulate healthy and aberrant mucosal healing following inflammation. In our previous funding period, as a necessary first step to towards achieving our long term goal, we developed a steady source of stable vocal fold epithelial cells to study epithelial cell-fibroblast interactions and established a novel physiologically relevant, three-dimensional biomimetic in vitro vocal fold model of human origin. For the proposed funding period, we will specifically focus on using a unique combination of in vitro and vivo model systems, including those developed in the previous funding period, to further our knowledge of host mediated immune defenses and inflammatory based tissue alterations in the vocal fold mucosa when exposed to defined bacteria species. The overall objective of this proposal is to identify mechanisms by which resident commensal or pathogen bacterial species are identified by vocal folds via specialized receptors and control tissue immune homeostasis while promoting immune responses in the context of steady state and tissue inflammation. In Specific Aim 1, we will define the role of commensal bacteria in tuning vocal fold immune cells using gnotobiotic methodologies. In Specific Aim 2, we will identify toll like family receptors (TLR) in vocal fold tissue and novel human vocal fold epithelial cells in steady state and inflammatory conditions. Lastly, in Specific Aim 3, using our physiologically relevant three- dimensional biomimetic model of normal and inflammatory vocal fold mucosa, we will determine relative contribution of resident commensal and pathogenic microbial communities on vocal fold mucosal integrity and function. Taken together, our multidisciplinary exploitation of microbiology, next gen sequencing, and immunology will converge to reform exponentially our understanding of mechanisms of innate immune- microbial interactions in vocal fold inflammatory diseases. 1